PROJECT SUMMARY To better investigate, identify and sense (detect) viruses, high-resolution separation, isolation and concentration is of significant importance. Our team is developing a bioparticle separation scheme based on dielectrophoresis using a microfluidic device that promises, and has demonstrated, extremely high-resolution separations of bioparticles. The separation is based on specific biophysical makeup of the viruses that in turn result in electrodynamic properties that vary for each specific target. If a separation scheme is of high enough resolution, distinct populations of homogeneous particles can be purified. Our preliminary data and recent theoretical modeling suggest that strain- or serotype-specific resolution of virus particles is achievable, however, this has not been explicitly shown. Unlabeled viruses must ultimately be used for the platform to be integrated in various virus assessment workflows. To enable detection of the separation of unlabeled virions, on-chip detection capabilities are developed, addressing this limiting issue in the current level of development. To demonstrate and develop the microfluidic platform we will use gene altered Sindbis virus (Venus and mCherry) which are fluorescent. Once parameters have been established for optimal dielectrophoresis in our system, the wild-type Sindbis virus (SINV) will be compared with mutants that harbor changes in the E2 protein to establish the ability to separate the viruses. Standard molecular biology genotyping and phenotyping will be used to confirm separation of virion populations. Figures of merit (detection limit, dynamic range, sensitivity and specificity) will be determined for the separated viruses along with defining appropriate properties of the incoming sample (ionic strength, pH, additives, etc.) to ensure proper operation of the technique.